Project Summary Alterations in serotonin (5HT) dynamics are observed in diverse clinical disorders including, but not limited to, stress and anxiety related syndromes. Interestingly, recent investigations have demonstrated an additional presence of ?reserve? pools of extravesicular 5HT in dorsal raphe (DRN) serotonergic neurons (both cytoplasmic and nuclear) however it has remained unclear whether it might play additional roles in the nucleus. A growing literature now demonstrates 5HT?s ability to form covalent bonds with certain cytoplasmic and membrane bound proteins via transamination by the tissue Transglutaminase 2 (TGM2) enzyme, a modification that has been suggested to alter the signaling properties of monoaminylated substrates. We therefore initially hypothesized that nuclear proteins in brain may similarly be modified to control distinct aspects of their function. Work from our group (Farrelly et al., Nature, 2018 ? re-review) has shown that 5HT can form covalent bonds with histone proteins (specifically histone H3; H3K4me3Q5ser)?a process known as serotonylation?which is critical for transcriptional permissiveness and early neuronal development. Thus, my plan during this K99/R00 proposal is to further explore the hypothesis that novel histone serotonylation may play critical roles in guiding normal patterns of transcriptional and cellular plasticity in brain during postnatal development and into adulthood, processes that may be disrupted in pathophysiological states associated with serotonergic dysfunction (i.e., chronic stress). Under the mentorship of Drs. Ian Maze and Li Shen at the Icahn School of Medicine at Mount Sinai New York, I will address this hypothesis in three distinct aims using a variety of approaches. During the K99 period (Aim 1) I plan to directly examine associations between H3K4me3Q5ser enrichment and transcriptional (dys)plasticity in developing and adult mouse brain (neurons vs. glia). This will also be investigated using a model of chronic social defeat stress (+/- antidepressants) as a mechanism to perturb serotonin signaling and thus further define a biological role for this modification in vivo. I will then assess (Aim 2) the contributions of H3 serotonylation to adulthood neural plasticity through genetic manipulations that alter levels of the mark, followed by extensive behavioral and transcriptional phenotyping. In my independent research program in Aim3 (R00), I will set out to further dissect distinct roles for H3Q5ser, as my preliminary data indicate that the serotonyl modification can exist in isolation of K4me3 and interacts uniquely in comparison to H3K4me3Q5ser with putative binding complexes (e.g., NuRD). This proposal promises to establish important patterns of transcriptional plasticity associated with histone serotonylation in brain. Moreover, the additional training afforded by this award and the scientific environment will ideally prepare me to launch an independent research program evaluating at multiple levels (epigenomic, cellular/behavioral) as to how histone serotonylation contributes to normal and abnormal states of plasticity.